Factors contributing to the development of chronic rejection in heterotopic rat heart transplantation

To monitor the low back risk imposed by asymmetric postures at workplaces, a method using angular velocity sensors was studied. According to a simple model analysis, trunk rotation could be calculated from the angular velocities measured at both the waist and shoulder and from the inclination of each angular velocity sensor. We thus developed a new detector consisting of an angular velocity sensor (ENC-05D, Murata, Japan) for detecting angular velocity and an acceleration sensor (ADXL05, Analog Devices, USA) for measuring inclination. The precision of the angular velocity sensor was high as the correlation coefficient between the output of the sensor and the true value was 0.9996. When the detectors were affixed to a subject and compared with data measured by a Vicon System 370 (Oxford Metrics, UK), the correlation coefficients between the two methods were 0.949 and 0.815 during model tasks of box transfer and box lifting, respectively. In a model of lifting boxes at different rates, the mean and standard deviation increased according to the task speed. This method was shown to be of practical use for monitoring trunk rotation.     

A humanized form of a CD4-specific monoclonal antibody exhibits decreased antigenicity and prolonged plasma half-life in rhesus monkeys while retaining its unique biological and antiviral properties

Certain monoclonal antibodies (MAbs) directed against CD4 can efficiently block HIV-1 replication in vitro. To explore CD4-directed passive immunotherapy for prevention or treatment of AIDS virus infection, we previously examined the biological activity of a nondepleting CD4-specific murine MAb, mu5A8. This MAb, specific for domain 2 of CD4, blocks HIV-1 replication at a post-gp120-CD4 binding step. When administered to normal rhesus monkeys, all CD4+ target cells were coated with antibody, yet no cell clearance or measurable immunosuppression occurred. However, strong anti-mouse Ig responses rapidly developed in all monkeys. In the present study, we report a successfully humanized form of mu5A8 (hu5A8) that retains binding to both human and monkey CD4 and anti-AIDS virus activity. When administered intravenously to normal rhesus monkeys, hu5A8 bound to all target CD4+ cells without depletion and showed a significantly longer plasma half-life than mu5A8. Nevertheless, an anti-hu5A8 response directed predominantly against V region determinants did eventually appear within 2 to 4 weeks in most animals. However, when hu5A8 was administered to rhesus monkeys chronically infected with the simian immunodeficiency virus of macaques, anti-hu5A8 antibodies were not detected. Repeated administration of hu5A8 in these animals resulted in sustained plasma levels and CD4+ cell coating with humanized antibody for 6 weeks. These studies demonstrate the feasibility of chronic administration of CD4-specific MAb as a potential means of treating or preventing HIV-1 infection.     

Rapid tyrosine phosphorylation of HS1 in the response of mouse lymphoma L5178Y-R cells to photodynamic treatment sensitized by the phthalocyanine Pc 4

The repeated administration of methamphetamine (METH) can result in long-lasting decreases in dopamine (DA) levels, tyrosine hydroxylase activity and DA uptake sites in the striatum. However, whether these changes lead to functional alterations in the dynamics of DA release and uptake has not been extensively examined. The present study used in vivo electrochemistry and microdialysis to examine potassium- and amphetamine-evoked release of DA in the striatum and nucleus accumbens (NAc) of METH-treated rats. Male Fischer-344 rats were administered METH (5 mg/kg s.c.) or saline four times in 1 day, at 2-hr intervals. One week later the animals were anesthetized with urethane and prepared for in vivo electrochemical recordings. The METH treatment resulted in dramatic decreases in potassium-evoked release of DA and in the rate of DA clearance in the striatum, whereas the NAc was not significantly affected. In vivo microdialysis studies demonstrated significant decreases in basal DA levels and in potassium- and amphetamine-evoked overflow of DA in the striatum of METH-treated animals. Basal and evoked DA levels in the NAc were not altered. Post-mortem levels of tissue DA were decreased by 41 to 67% in the striatum and 25 to 31% in the NAc. These results indicate that the striatum is more sensitive than the NAc to the neurotoxic effects of METH, both in measures of functional dynamics of DA signaling and in tissue levels of DA. It remains to be determined whether these functional changes in DA release and uptake are permanent or tend to recover over time.     

Binding of tsHMG, a mouse testis-specific HMG-domain protein, to cisplatin-DNA adducts

The anticancer drug cisplatin is particularly effective against testicular tumors. Although the clinical consequences of cisplatin chemotherapy are well-known, the precise mechanism of action remains elusive. Specific recognition of cisplatin-damaged DNA by a class of proteins containing the high-mobility group (HMG) domain DNA-binding motif could play a role in mediating the cytotoxicity of the drug. This study presents a quantitative investigation of binding of the murine testis-specific high-mobility group protein tsHMG to DNA modified by cisplatin. The binding affinity and specificity of this protein to a site-specific 1,2-d(GpG) cisplatin-DNA intrastrand cross-link in a 20 bp probe were determined. A value for the apparent dissociation constant, Kd(app), of 24 +/- 5 nM was obtained by gel mobility shift assays. Binding competition assays with the corresponding unmodified 20 bp probe gave a ratio (rho) of nonspecific to specific Kd(app) values of 230. A polypeptide containing tsHMG domain A (residues 1-82) was expressed and purified to homogeneity. This domain alone was sufficient for specific recognition of cisplatin-modified DNA with a Kd(app) of 300 +/- 50 nM and a rho of 20, a comparatively high discrimination factor. DNase I interference analysis of the adduct-containing strand revealed that tsHMG binding extends over 14 nucleotides, centered around the platinated bases. The domain A polypeptide protection pattern covers a slightly smaller area of 13 nucleotides. The binding affinity and specificity of tsHMG for cisplatin-modified DNA are exceptional compared to those of other HMG-domain proteins studied previously. The possible relevance of these findings to the mechanism of action of cisplatin is discussed.     

Characterization of a human papillomavirus type 16 variant-dependent neutralizing epitope

We have determined that three type-specific and conformationally dependent monoclonal antibodies, H16.E70, H16.U4, and H16.V5, neutralize pseudotype human papillomavirus type 16 (HPV16) virions in vitro. H16.U4 and H16.V5 neutralized pseudotype virions derived from the German HPV16 variant 114K and the Zairian variant Z-1194 with equal efficiency. In contrast, neutralization of Z-1194 pseudotype virions by H16.E70 was two orders of magnitude weaker than neutralization of 114K pseudotype virions. This difference correlated with enzyme-linked immunosorbent assay reactivity of H16.E70 to L1 virus-like particles of the two variants. A substitution at residue 282 of L1 was responsible for this differential reactivity, suggesting that this residue constitutes part of the H16.E70 epitope.     

Cyclic guanosine monophosphate responses to atrial natriuretic factor, brain natriuretic peptide, but not C-type natriuretic peptide, and the characterization of their receptors in rat medullary thick ascending limb

The effects of atrial natriuretic factor (ANF), brain natriuretic peptide (BNP), and C-type natriuretic peptide (CNP) on renal medullary thick ascending limb (mTAL) have not been fully understood. The aim of this study is to examine the second-messenger responses of rat mTAL to ANF, BNP, and CNP. Characterizations of the ANF, BNP, and CNP receptors in mTAL were also performed by radioligand studies. Results showed that ANF and BNP were both capable of eliciting cyclic guanosine monophosphate (cGMP) responses in mTAL. Conversely, no cGMP response was observed upon stimulation by CNP in mTAL. The presence of ANF receptors was demonstrated by radioligand studies. One receptor site was found, and the Kd and maximum binding capacity were 4.0 +/- 0.45 nmol/L and 277.8 +/- 47.7 fmol/mg protein, respectively. BNP receptors were also found in mTAL, and ANF and BNP were sharing the same receptor. On the contrary, no CNP receptor could be shown by radioligand studies. These results suggest that guanylyl cyclase-coupled receptors (atrial natriuretic peptide receptor-A [ANPR-A]) specific for ANF and BNP are present in rat mTAL, while those for CNP (ANPR-B) are absent. ANF and BNP but not CNP act on mTAL to control water excretion.     

The light chain of CD98 is identified as E16/TA1 protein

The 80/40-kDa CD98 protein complex was purified using an anti-CD98 heavy chain monoclonal antibody coupled to Sepharose beads. Eluted proteins were subjected to preparative SDS-polyacrylamide gel electrophoresis, and protein corresponding to the 40-kDa CD98 light chain was excised. Following proteolysis with trypsin, a peptide fragment was sequenced by mass spectrometry. The nine residues obtained were identical to established C-terminal sequences of the human E16 and rat TA1 proteins, suggesting that TA1/E16 protein is the CD98 light chain. Consistent with this, anti-TA1/E16 antibodies specifically immunoblotted the approximately 35-40-kDa light chain present upon immunoprecipitation of the human CD98 complex. Furthermore, anti-CD98 heavy chain antibody specifically co-immunoprecipitated hemagglutinin-tagged light chain from cells transfected with hemagglutinin-tagged E16 cDNA. In conclusion, the CD98 light chain is identical to the TA1/E16 protein, based on partial amino acid sequence identity, antibody cross-reactivity, genetic reconstitution evidence, similar molecular size, and comparable cell distribution.     

TGF-beta: a balancing act

Regulation of developmental processes as well as host defense and repair mechanisms requires the maintenance of a delicate balance of positive and negative regulatory signals. TGF-beta, a molecule known for its many diverse activities, can promote or inhibit cell growth and function. Disruption of the balance between these opposing activities can contribute to aberrant development, malignancy, or pathogenic immune and inflammatory responses. TGF-beta transgenic mouse studies highlight the essential function(s) of TGF-beta and its receptors and provide insight to potential therapeutic approaches to manipulate TGF-beta expression.